The long-term goal of the laboratory will involve a complete biochemical and genetic analysis of the mechanism and control of fatty acid movement into and out of adipose tissue. The flux of fatty acids through triglyceride is a hormonally controlled process; influx or synthesis dependent upon insulin, effelux upon epinephrine. The goal of the laboratory will center around identification of the mechanism(s) by which these hormones control adipocyte triglyceride levels. Additionaly, the expression of genes encoding the adipocyte fatty acid carrier and fatty acid transporter are conjectured to also be under hormonal influences such that the circulating levels of insulin and epinephrine may regulate gene expression, mRNA stability, and protein function. The initial studies of the laboratory will center upon a complete characterization of the adipocyte fatty acid carrier. To do this the 3T3-L1 differentiating preadipocyte cell line appears to be an ideal model system for several reasons: 1) 3T3-L1 adipocytes respond to lipolytic and lipogenic hormones, 2) a full length cDNA clone for the putative fatty acid carrier has been isolated, and 3) the differentiation-dependent expression of the fatty acid carrier gene can be dissected and its hormonal control examined. To accomplish this the following will be undertaken: A. Purification of the adipocyte-specific fatty acid carrier. B. Biochemical studies of the affinity and structure of the bound fatty acid. C. Production of monoclonal antibodies specific to the carrier to measure the effect of insulin on the proteins abundance and intracellular location. D. Measure the effect of insulin on the mRNA abundance and gene expression by nuclear runoff transcription E. Clone and characterize the structural gene for the carrier. F. Identify by transient expression and stable transformation the DNA sequences regulating expression during differentiation and regulation by insulin. G. Measure by nuclease sensitivity the alterations in chromatin structure which accompanies gene expression. The results of such an investigation will hopefully provide insights into the biochemical mechanisms underlying the clinical problems such as chronic obesity, diabetic obesity and human hyperchylomicronemias for these conditions undoubtedly involve, to some extent, mishandling of fatty acids in adipose tissue.